Nitrate is a key required nutrient for the synthesis of amino acids, nucleotides and vitamins and is commonly considered to be the most limiting for normal plant growth (Vitousek et al., 2004, Biogeochemistry 13). Nitrogenous fertilizer is usually supplied as ammonium nitrate, potassium nitrate, or urea. Plants are keenly sensitive to nitrogen levels in the soil and, atypically of animal development, adopt their body plan to cope with their environment (Lopez-Bucio et al., 2003, Curr Opin Plant Biol 6:280-287; Malamy et al., 2005, Plant Cell Environ 28:67-77; Walch-Liu et al., 2006, Ann Bot (Lond) 97:875-81). For example, mutants in several general nitrogen (N)-assimilation genes affect root architecture (Little et al., 2005, Proc Natl Acad Sci USA 102:13693-13698; Remans et al., 2006, Proc Natl Acad Sci USA 103:19206-19211). Transduction of this nitrogen signal is linked to a massive and concerted gene expression response in the root (Gutierrez et al., 2007, Genome Biol 8:R7; Wang et al., 2003, Plant Physiol 132:556-67).
Studies on the regulation of genes involved in the N-assimilatory pathway have shown that genes involved in N-assimilation are regulated transcriptionally by both inorganic and organic forms of nitrogen (FIG. 1). Genes involved in the uptake and reduction of nitrate (NIA, NIR) are transcriptionally induced by nitrate. By contrast, the glutamine synthetase gene (GLN1.3) involved in assimilating inorganic N into organic form (Gln), is transcriptionally repressed by the endproducts of N-assimilation (Glu/Gln) (FIG. 1). The repression of GLN1.3 expression by the product of the GS enzyme reaction serves as a negative feedback loop, that shuts off further assimilation of inorganic N into Gln, when levels of Gln are abundant. As GS is and ATP dependent enzyme, this is likely to be an energy conservation mechanism. By contrast, Gln/Glu levels activate the expression of the ASN1 gene (asparagine synthetase) which serves to transfer the amide N from Gln onto Asp to make Asn and Glu as a by-product. Asn is an inert amino acid used to store N and used for long distance N-transport (e.g., to seed). The induction of ASN1 by Glu/Gln is a mechanism that serves to store excess N as Asn, which is used to transport N to seed.
It would be advantageous to produce plants that would continue to assimilate and utilize N or to store N depending on whether a vegetative plant part or seed is the product, respectively, making N-assimilation independent of the Glu/Gln biofeedback pathway. N-assimilated into Glu/Gln by GS is used in the biosynthesis of all N-containing compounds including essentially all other amino acids, nucleic acids and chlorophyll. By contrast, the conversion of Gln to Asn (an inert N source) is used to transport and store N in seed.